The success of vaccines depends upon the induction of strong, long lasting immune responses that will protect the individual from a subsequent encounter with the infectious agent. Epidemiologic evidence and results of clinical trials suggests that the development of an effective vaccine against Shigella is an achievable goal. Adult volunteers who were experimentally infected with either wild type S. sonnei or S. flexneri 2a at the CVD were significantly protected against illness following rechallenge with the homologous strain. However, the presence of antibodies to Shigella measured in these volunteers did not correlate with resistance to shigellosis following challenge. Cell-mediated immunity (CMI) is a key effector mechanism involved in protection against many intracellular pathogens, including bacteria. However, very limited information is available concerning the role of CMI in Shigella infection. The overall goal of the proposed studies is to identify the immunological mechanisms that mediate effective protection from shigellosis following vaccination and natural infection with Shigella. Our hypothesis is that CMI responses play a central role in protection of volunteers from shigellosis following infection with wild-type Shigella and live Shigella vaccine candidates. The identification of CMI that correlate with protection following exposure to wild-type Shigella will greatly enhance our understanding of the mechanisms underlying protection in shigellosis and the development of Shigella vaccines. Specifically, using cells isolated from mucosal biopsies and peripheral blood mononuclear cells (PBMC) obtained from volunteers immunized with wild-type or attenuated strains of S. flexneri 2a, S. dysenteriae and S. sonnei we propose to test the following hypotheses: (1) Challenge with wild-type Shigella or immunization with Shigella vaccine candidates elicits the appearance in circulation of specific cytotoxic T lymphocytes (CTL); (2) Challenge with wild-type Shigella or immunization with attenuated strains of Shigella elicits the appearance in circulation of specific T lymphocytes that proliferate and produce interferon-gamma interferon-gamma (IFN-gamma), as well as other "type-1" cytokines, in response to Shigella antigens, (3) Challenge with wild-type Shigella or immunization with attenuated strains of Shigella elicits the appearance in the gut mucosa of specific CTL effectors and T lymphocytes that proliferate and produce IFN- gamma, as well as other "type-1" cytokines, in response to Shigella antigens; (4) CTL responses and/or IFN-gamma production elicited by immunization at the mucosal and systemic levels correlate with protection to challenge with virulent Shigella strains; and (5) Determine whether "protective epitopes" can be identified in Shigella antigens.